Myofibroblast-specific inhibition of the Rho kinase-MRTF-SRF pathway using nanotechnology for the prevention of pulmonary fibrosis.

Pulmonary fibrosis is characterized by the accumulation of myofibroblasts in the lung and progressive tissue scarring. Fibroblasts exist across a spectrum of states, from quiescence in health to activated myofibroblasts in the setting of injury. Highly activated myofibroblasts have a critical role in the establishment of fibrosis as the predominant source of type 1 collagen and profibrotic mediators. Myofibroblasts are also highly contractile cells and can alter lung biomechanical properties through tissue contraction. Inhibiting signaling pathways involved in myofibroblast activation could therefore have significant therapeutic value. One of the ways myofibroblast activation occurs is through activation of the Rho/myocardin-related transcription factor (MRTF)/serum response factor (SRF) pathway, which signals through intracellular actin polymerization. However, concerns surrounding the pleiotropic and ubiquitous nature of these signaling pathways have limited the translation of inhibitory drugs. Herein, we demonstrate a novel therapeutic antifibrotic strategy using myofibroblast-targeted nanoparticles containing a MTRF/SRF pathway inhibitor (CCG-1423), which has been shown to block myofibroblast activation in vitro. Myofibroblasts were preferentially targeted via the angiotensin 2 receptor, which has been shown to be selectively upregulated in animal and human studies. These nanoparticles were nontoxic and accumulated in lung myofibroblasts in the bleomycin-induced mouse model of pulmonary fibrosis, reducing the number of these activated cells and their production of profibrotic mediators. Ultimately, in a murine model of lung fibrosis, a single injection of these drugs containing targeted nanoagents reduced fibrosis as compared with control mice. This approach has the potential to deliver personalized therapy by precisely targeting signaling pathways in a cell-specific manner, allowing increased efficacy with reduced deleterious off-target effects.

Multisensory contribution in visuospatial orientation: an interaction between neck and trunk proprioception.

A coherent perception of spatial orientation is key in maintaining postural control. To achieve this the brain must access sensory inputs encoding both the body and the head position and integrate them with incoming visual information. Here we isolated the contribution of proprioception to verticality perception and further investigated whether changing the body position without moving the head can modulate visual dependence-the extent to which an individual relies on visual cues for spatial orientation. Spatial orientation was measured in ten healthy individuals [6 female; 25-47 years (SD 7.8 years)] using a virtual reality based subjective visual vertical (SVV) task. Individuals aligned an arrow to their perceived gravitational vertical, initially against a static black background (10 trials), and then in other conditions with clockwise and counterclockwise background rotations (each 10 trials). In all conditions, subjects were seated first in the upright position, then with trunk tilted 20° to the right, followed by 20° to the left while the head was always aligned vertically. The SVV error was modulated by the trunk position, and it was greater when the trunk was tilted to the left compared to right or upright trunk positions (p < 0.001). Likewise, background rotation had an effect on SVV errors as these were greater with counterclockwise visual rotation compared to static background and clockwise roll motion (p < 0.001). Our results show that the interaction between neck and trunk proprioception can modulate how visual inputs affect spatial orientation.

Atorvastatin Reduces In Vivo Fibrin Deposition and Macrophage Accumulation, and Improves Primary Patency Duration and Maturation of Murine Arteriovenous Fistula.

BACKGROUND: Arteriovenous fistulas placed surgically for dialysis vascular access have a high primary failure rate resulting from excessive inward remodeling, medial fibrosis, and thrombosis. No clinically established pharmacologic or perisurgical therapies currently address this unmet need. Statins' induction of multiple anti-inflammatory and antithrombotic effects suggests that these drugs might reduce arteriovenous fistula failure. Yet, the in vivo physiologic and molecular effects of statins on fistula patency and maturation remain poorly understood. METHODS: We randomized 108 C57Bl/6J mice to receive daily atorvastatin 1.14 mg/kg or PBS (control) starting 7 days before end-to-side carotid artery-jugular vein fistula creation and for up to 42 days after fistula creation. We then assessed longitudinally the effects of statin therapy on primary murine fistula patency and maturation. We concomitantly analyzed the in vivo arteriovenous fistula thrombogenic and inflammatory macrophage response to statin therapy, using the fibrin-targeted, near-infrared fluorescence molecular imaging agent FTP11-CyAm7 and dextranated, macrophage-avid nanoparticles CLIO-VT680. RESULTS: In vivo molecular-structural imaging demonstrated that atorvastatin significantly reduced fibrin deposition at day 7 and macrophage accumulation at days 7 and 14, findings supported by histopathologic and gene-expression analyses. Structurally, atorvastatin promoted favorable venous limb outward remodeling, preserved arteriovenous fistula blood flow, and prolonged primary arteriovenous fistula patency through day 42 (P<0.05 versus control for all measures). CONCLUSIONS: These findings provide new in vivo evidence that statins improve experimental arteriovenous fistula patency and maturation, indicating that additional clinical evaluation of statin therapy in patients on dialysis undergoing arteriovenous fistula placement is warranted.

Vigorous cool room treadmill training to improve walking ability in people with multiple sclerosis who use ambulatory assistive devices: a feasibility study.

BACKGROUND: Aerobic training has the potential to restore function, stimulate brain repair, and reduce inflammation in people with Multiple Sclerosis (MS). However, disability, fatigue, and heat sensitivity are major barriers to exercise for people with MS. We aimed to determine the feasibility of conducting vigorous harness-supported treadmill training in a room cooled to 16 °C (10 weeks; 3times/week) and examine the longer-term effects on markers of function, brain repair, and inflammation among those using ambulatory aids. METHODS: Ten participants (9 females) aged 29 to 74 years with an Expanded Disability Status Scale ranging from 6 to 7 underwent training (40 to 65% heart rate reserve) starting at 80% self-selected walking speed. Feasibility of conducting vigorous training was assessed using a checklist, which included attendance rates, number of missed appointments, reasons for not attending, adverse events, safety hazards during training, reasons for dropout, tolerance to training load, subjective reporting of symptom worsening during and after exercise, and physiological responses to exercise. Functional outcomes were assessed before, after, and 3 months after training. Walking ability was measured using Timed 25 Foot Walk test and on an instrumented walkway at both fast and self-selected speeds. Fatigue was measured using fatigue/energy/vitality sub-scale of 36-Item Short-Form (SF-36) Health Survey, Fatigue Severity Scale, modified Fatigue Impact Scale. Aerobic fitness (maximal oxygen consumption) was measured using maximal graded exercise test (GXT). Quality-of-life was measured using SF-36 Health Survey. Serum levels of neurotrophin (brain-derived neurotrophic factor) and cytokine (interleukin-6) were assessed before and after GXT. RESULTS: Eight of the ten participants completed training (attendance rates ≥ 80%). No adverse events were observed. Fast walking speed (cm/s), gait quality (double-support (%)) while walking at self-selected speed, fatigue (modified Fatigue Impact Scale), fitness (maximal workload achieved during GXT), and quality-of-life (physical functioning sub-scale of SF-36) improved significantly after training, and improvements were sustained after 3-months. Improvements in fitness (maximal respiratory exchange ratio and maximal oxygen consumption during GXT) were associated with increased brain-derived neurotrophic factor and decreased interleukin-6. CONCLUSION: Vigorous cool room training is feasible and can potentially improve walking, fatigue, fitness, and quality-of-life among people with moderate to severe MS-related disability. TRIAL REGISTRATION: The study was approved by the Newfoundland and Labrador Health Research Ethics Board (reference number: 2018.088) on 11/07/2018 prior to the enrollment of first participant (retrospectively registered at ClinicalTrials.gov: NCT04066972. Registered on 26 August 2019.

Intravascular fibrin molecular imaging improves the detection of unhealed stents assessed by optical coherence tomography in vivo.

AIMS: Fibrin deposition and absent endothelium characterize unhealed stents that are at heightened risk of stent thrombosis. Optical coherence tomography (OCT) is increasingly used for assessing stent tissue coverage as a measure of healed stents, but cannot precisely identify whether overlying tissue represents physiological neointima. Here we assessed and compared fibrin deposition and persistence on bare metal stent (BMS) and drug-eluting stent (DES) using near-infrared fluorescence (NIRF) molecular imaging in vivo, in combination with simultaneous OCT stent coverage. METHODS AND RESULTS: Rabbits underwent implantation of one BMS and one DES without overlap in the infrarenal aorta (N = 20 3.5 × 12 mm). At Days 7 and/or 28, intravascular NIRF-OCT was performed following the injection of fibrin-targeted NIRF molecular imaging agent FTP11-CyAm7. Intravascular NIRF-OCT enabled high-resolution imaging of fibrin overlying stent struts in vivo, as validated by histopathology. Compared with BMS, DES showed greater fibrin deposition and fibrin persistence at Days 7 and 28 (P < 0.01 vs. BMS). Notably, for edge stent struts identified as covered by OCT on Day 7, 92.8 ± 9.5% of DES and 55.8 ± 23.6% of BMS struts were NIRF fibrin positive (P < 0.001). At Day 28, 18.6 ± 10.6% (DES) and 5.1 ± 8.7% (BMS) of OCT-covered struts remained fibrin positive (P < 0.001). CONCLUSION: Intravascular NIRF fibrin molecular imaging improves the detection of unhealed stents, using clinically translatable technology that complements OCT. A sizeable percentage of struts deemed covered by OCT are actually covered by fibrin, particularly in DES, and therefore such stents might remain prothrombotic. These findings have implications for the specificity of standalone clinical OCT assessments of stent healing.

Bone Fracture Acute Phase Response-A Unifying Theory of Fracture Repair: Clinical and Scientific Implications.

Bone fractures create five problems that must be resolved: bleeding, risk of infection, hypoxia, disproportionate strain, and inability to bear weight. There have been enormous advancements in our understanding of the molecular mechanisms that resolve these problems after fractures, and in best clinical practices of repairing fractures. We put forth a modern, comprehensive model of fracture repair that synthesizes the literature on the biology and biomechanics of fracture repair to address the primary problems of fractures. This updated model is a framework for both fracture management and future studies aimed at understanding and treating this complex process. This model is based upon the fracture acute phase response (APR), which encompasses the molecular mechanisms that respond to injury. The APR is divided into sequential stages of "survival" and "repair." Early in convalescence, during "survival," bleeding and infection are resolved by collaborative efforts of the hemostatic and inflammatory pathways. Later, in "repair," avascular and biomechanically insufficient bone is replaced by a variable combination of intramembranous and endochondral ossification. Progression to repair cannot occur until survival has been ensured. A disproportionate APR-either insufficient or exuberant-leads to complications of survival (hemorrhage, thrombosis, systemic inflammatory response syndrome, infection, death) and/or repair (delayed- or non-union). The type of ossification utilized for fracture repair is dependent on the relative amounts of strain and vascularity in the fracture microenvironment, but any failure along this process can disrupt or delay fracture healing and result in a similar non-union. Therefore, incomplete understanding of the principles herein can result in mismanagement of fracture care or application of hardware that interferes with fracture repair. This unifying model of fracture repair not only informs clinicians how their interventions fit within the framework of normal biological healing but also instructs investigators about the critical variables and outputs to assess during a study of fracture repair.

Imaging Granzyme B Activity Assesses Immune-Mediated Myocarditis.

RATIONALE: The development of molecular imaging approaches that assess specific immunopathologic mechanisms can advance the study of myocarditis. OBJECTIVE: This study validates a novel molecular imaging tool that enables the in vivo visualization of granzyme B activity, a major effector of cytotoxic CD8+ T lymphocytes. METHODS AND RESULTS: We synthesized and optimized a fluorogenic substrate capable of reporting on granzyme B activity and examined its specificity ex vivo in mice hearts with experimental cytotoxic CD8+ T lymphocyte-mediated myocarditis using fluorescence reflectance imaging, validated by histological examination. In vivo experiments localized granzyme B activity in hearts with acute myocarditis monitored by fluorescent molecular tomography in conjunction with coregistered computed tomography imaging. A model anti-inflammatory intervention (dexamethasone administration) in vivo reduced granzyme B activity (vehicle versus dexamethasone: 504±263 versus 194±77 fluorescence intensities in hearts; P=0.002). CONCLUSIONS: Molecular imaging of granzyme B activity can visualize T cell-mediated myocardial injury and monitor the response to an anti-inflammatory intervention.

In vivo nanoparticle assessment of pathological endothelium predicts the development of inflow stenosis in murine arteriovenous fistula.

OBJECTIVE: In vivo assessment of pathological endothelium within arteriovenous fistula (AVF) could provide new insights into inflow stenosis, a common cause of AVF primary failure in end-stage renal disease patients. Here we developed nanoparticle-based imaging strategies to assess pathological endothelium in vivo and elucidate its relationship to neointimal hyperplasia formation in AVF. APPROACH AND RESULTS: Jugular-carotid AVFs were created in C57BL/6 mice (n=38). Pathological endothelium in the AVF was visualized and quantified in vivo using dextranated magnetofluorescent nanoparticles (CLIO-VT680 [cross-linked iron oxide-VivoTag680]). At day 14, CLIO-VT680 was deposited in AVF, but only minimally in sham-operated arteries. Transmission electron microscopy revealed that CLIO-VT680 resided within endothelial cells and in the intimal extracellular space. Endothelial cells of AVF, but not control arteries, expressed vascular cell adhesion molecule-1 and showed augmented endothelial permeability near the anastomosis. Intravital microscopy demonstrated that CLIO-VT680 deposited most intensely near the AVF anastomosis (P<0.0001). The day 14 intravital microscopy CLIO-VT680 signal predicted the subsequent site and magnitude of AVF neointimal hyperplasia at day 42 (r=0.58, P<0.05). CLIO-VT680 deposition in AVF was further visualized by ex vivo MRI. CONCLUSIONS: AVF develop a pathological endothelial response that can be assessed in vivo via nanoparticle-enhanced imaging. AVF endothelium is activated and exhibits augmented permeability, offering a targeting mechanism for nanoparticle deposition and retention in pathological endothelium. The in vivo AVF nanoparticle signal identified and predicted subsequent inflow neointimal hyperplasia. This approach could be used to test therapeutic interventions aiming to restore endothelial health and to decrease early AVF failure caused by inflow stenosis.

Diagnosis and Management of Vertebral Compression Fractures.

Vertebral compression fractures (VCFs) are the most common complication of osteoporosis, affecting more than 700,000 Americans annually. Fracture risk increases with age, with four in 10 white women older than 50 years experiencing a hip, spine, or vertebral fracture in their lifetime. VCFs can lead to chronic pain, disfigurement, height loss, impaired activities of daily living, increased risk of pressure sores, pneumonia, and psychological distress. Patients with an acute VCF may report abrupt onset of back pain with position changes, coughing, sneezing, or lifting. Physical examination findings are often normal, but can demonstrate kyphosis and midline spine tenderness. More than two-thirds of patients are asymptomatic and diagnosed incidentally on plain radiography. Acute VCFs may be treated with analgesics such as acetaminophen, nonsteroidal anti-inflammatory drugs, narcotics, and calcitonin. Physicians must be mindful of medication adverse effects in older patients. Other conservative therapeutic options include limited bed rest, bracing, physical therapy, nerve root blocks, and epidural injections. Percutaneous vertebral augmentation, including vertebroplasty and kyphoplasty, is controversial, but can be considered in patients with inadequate pain relief with nonsurgical care or when persistent pain substantially affects quality of life. Family physicians can help prevent vertebral fractures through management of risk factors and the treatment of osteoporosis.

Biomimetic Nanomaterials for the Immunomodulation of the Cardiosplenic Axis Postmyocardial Infarction.

The spleen is an important mediator of both adaptive and innate immunity. As such, attempts to modulate the immune response provided by the spleen may be conducive to improved outcomes for numerous diseases throughout the body. Here, biomimicry is used to rationally design nanomaterials capable of splenic retention and immunomodulation for the treatment of disease in a distant organ, the postinfarct heart. Engineered senescent erythrocyte-derived nanotheranostic (eSENTs) are generated, demonstrating significant uptake by the immune cells of the spleen including T and B cells, as well as monocytes and macrophages. When loaded with suberoylanilide hydroxamic acid (SAHA), the nanoagents exhibit a potent therapeutic effect, reducing infarct size by 14% at 72 h postmyocardial infarction when given as a single intravenous dose 2 h after injury. These results are supportive of the hypothesis that RBC-derived biomimicry may provide new approaches for the targeted modulation of the pathological processes involved in myocardial infarction, thus further experiments to decisively confirm the mechanisms of action are currently underway. This novel concept may have far-reaching applicability for the treatment of a number of both acute and chronic conditions where the immune responses are either stimulated or suppressed by the splenic (auto)immune milieu.

Ultrathin-strut versus thin-strut stent healing and outcomes in preclinical and clinical subjects.

BACKGROUND: Compared with thin-strut durable-polymer drug-eluting stents (DP-DES), ultrathin-strut biodegradable-polymer sirolimus-eluting stents (BP-SES) improve stent-related clinical outcomes in patients undergoing percutaneous coronary intervention (PCI). Reduced stent strut thickness is hypothesised to underlie these benefits, but this conjecture remains unproven. AIMS: We aimed to assess the impact of strut thickness on stent healing and clinical outcomes between ultrathin-strut and thin-strut BP-SES. METHODS: First, we performed a preclinical study of 8 rabbits implanted with non-overlapping thin-strut (diameter/thickness 3.5 mm/80 µm) and ultrathin-strut (diameter/thickness 3.0 mm/60 µm) BP-SES in the infrarenal aorta. On day 7, the rabbits underwent intravascular near-infrared fluorescence optical coherence tomography (NIRF-OCT) molecular-structural imaging of fibrin deposition and stent tissue coverage, followed by histopathological analysis. Second, we conducted an individual data pooled analysis of patients enrolled in the BIOSCIENCE and BIOSTEMI randomised PCI trials treated with ultrathin-strut (n=282) or thin-strut (n=222) BP-SES. The primary endpoint was target lesion failure (TLF) at 1-year follow-up, with a landmark analysis at 30 days. RESULTS: NIRF-OCT image analyses revealed that ultrathin-strut and thin-strut BP-SES exhibited similar stent fibrin deposition (p=0.49) and percentage of uncovered stent struts (p=0.63). Histopathological assessments corroÂborated these findings. In 504 pooled randomised trial patients, TLF rates were similar for those treated with ultrathin-strut or thin-strut BP-SES at 30-day (2.5% vs 1.8%; p=0.62) and 1-year follow-up (4.3% vs 4.7%; p=0.88). CONCLUSIONS: Ultrathin-strut and thin-strut BP-SES demonstrate similar early arterial healing profiles and 30-day and 1-year clinical outcomes.

Inflammation modulates murine venous thrombosis resolution in vivo: assessment by multimodal fluorescence molecular imaging.

OBJECTIVE: Assessment of thrombus inflammation in vivo could provide new insights into deep vein thrombosis (DVT) resolution. Here, we develop and evaluate 2 integrated fluorescence molecular-structural imaging strategies to quantify DVT-related inflammation and architecture and to assess the effect of thrombus inflammation on subsequent DVT resolution in vivo. METHODS AND RESULTS: Murine DVT were created with topical 5% FeCl(3) application to thigh or jugular veins (n=35). On day 3, mice received macrophage and matrix metalloproteinase activity fluorescence imaging agents. On day 4, integrated assessment of DVT inflammation and architecture was performed using confocal fluorescence intravital microscopy. Day 4 analyses showed robust relationships among in vivo thrombus macrophages, matrix metalloproteinase activity, and fluorescein isothiocyanate-dextran deposition (r>0.70; P<0.01). In a serial 2-time point study, mice with DVT underwent intravital microscopy at day 4 and day 6. Analyses revealed that the intensity of thrombus inflammation at day 4 predicted the magnitude of DVT resolution at day 6 (P<0.05). In a second approach, noninvasive fluorescence molecular tomography-computed tomography was used and detected macrophages within jugular DVT (P<0.05 versus sham controls). CONCLUSIONS: Integrated fluorescence molecular-structural imaging demonstrates that the DVT-induced inflammatory response can be readily assessed in vivo and can inform the magnitude of thrombus resolution.

Polymeric nanomaterials for islet targeting and immunotherapeutic delivery.

Here we report a proof-of-concept for development of pancreatic islet-targeting nanoparticles for immunomodulatory therapy of autoimmune type 1 diabetes. Modified with a unique islet-homing peptide, these polymeric nanomaterials exhibit 3-fold greater binding to islet endothelial cells and a 200-fold greater anti-inflammatory effect through targeted islet endothelial cell delivery of an immunosuppressant drug. Our findings also underscore the need to carefully tailor drug loading and nanoparticle dosage to achieve maximal vascular targeting and immunosuppression.

The implementation of a precision case management model in a Canadian inpatient rehabilitation center: The 12-months post-implementation findings of a quality improvement project.

Despite recommendations, few have reported on quality improvement initiatives to implement length of rehabilitation stay benchmarks, while actively monitoring functional outcomes. This article describes the development, implementation, and evaluation of a precision case management model across all inpatient rehabilitation client groups in a Canadian facility. To develop the length of rehabilitation-stay (LoRS) benchmarks, patient data was retrospectively analyzed. A severity specific method was used to stratify median length of stay. A target reduction on 8.6 days in LoRS was established. Functional discharge targets were also set and monitored at specific intervals via the Functional Independence Measure (FIM®). The implementation used an incremental quality improvement phased approach. Following 12-months, a statistically significant reduction in mean LoRS of 13.2 days was achieved, along with a small increase in FIM® change across all rehabilitation client groups. A similar pattern was seen across the three main client groups, where a LoRS reduction greater than the target was achieved, along with important improvements in LoRS efficiency. This study demonstrates how the implementation of a precision case management model can assist a facility in markedly reducing LoRS across inpatient groups, without compromising functional change or community discharge rates and begin its transformation to a value-based organization.

meso-arylporpholactones and their reduction products.

The rational syntheses of meso-tetraaryl-3-oxo-2-oxaporphyrins 5, known as porpholactones, via MnO(4)(-)-mediated oxidations of the corresponding meso-tetraaryl-2,3-dihydroxychlorins (7) is detailed. Since chlorin 7 is prepared from the parent porphyrin 1, this amounts to a 2-step replacement of a pyrrole moiety in 1 by an oxazolone moiety. The stepwise reduction of the porpholactone 5 results in the formation of chlorin analogues, meso-tetraaryl-3-hydroxy-2-oxachlorin (11) and meso-tetraaryl-2-oxachlorins (12). The reactivity of 11 with respect to nucleophilic substitution by O-, N-, and S-nucleophiles is described. The profound photophysical consequences of the formal replacement of a pyrrole with an oxazolone (porphyrin-like chromophore) or (substituted) oxazole moiety (chlorin-like chromophore with, for the parent oxazolochlorin 12, red-shifted Q(x) band with enhanced oscillator strengths) are detailed and rationalized on the basis of SAC-CI and MNDO-PSDCI molecular orbital theory calculations. The single crystal X-ray structures of the porpholactones point at a minor steric interaction between the carbonyl oxygen and the flanking phenyl group. The essentially planar structures of all chromophores in all oxidation states prove that the observed optical properties originate from the intrinsic electronic properties of the chromophores and are not subject to conformational modulation.

Engineered Cell-Derived Vesicles Displaying Targeting Peptide and Functionalized with Nanocarriers for Therapeutic microRNA Delivery to Triple-Negative Breast Cancer in Mice.

Polymeric nanocarriers (PNCs) can be used to deliver therapeutic microRNAs (miRNAs) to solid cancers. However, the ability of these nanocarriers to specifically target tumors remains a challenge. Alternatively, extracellular vesicles (EVs) derived from tumor cells show homotypic affinity to parent cells, but loading sufficient amounts of miRNAs into EVs is difficult. Here, it is investigated whether uPAR-targeted delivery of nanococktails containing PNCs loaded with therapeutic antimiRNAs, and coated with uPA engineered extracellular vesicles (uPA-eEVs) can elicit synergistic antitumor responses. The uPA-eEVs coating on PNCs increases natural tumor targeting affinities, thereby enhancing the antitumor activity of antimiRNA nanococktails. The systemic administration of uPA-eEV-PNCs nanococktail shows a robust tumor tropism, which significantly enhances the combinational antitumor effects of antimiRNA-21 and antimiRNA-10b, and leads to significant tumor regression and extension of progression free survival for syngeneic 4T1 tumor-bearing mice. In addition, the uPA-eEV-PNCs-antimiRNAs nanococktail plus low dose doxorubicin results in a synergistic antitumor effect as evidenced by inhibition of tumor growth, reduction of lung metastases, and extension of survival of 4T1 tumor-bearing mice. The targeted combinational nanococktail strategy could be readily translated to the clinical setting by using autologous cancer cells that have flexibility for ex vivo expansion and genetic engineering.

Helimeric porphyrinoids: stereostructure and chiral resolution of meso-tetraarylmorpholinochlorins.

The synthesis and chiral resolution of free-base and Ni(II) complexes of a number of derivatives of meso-tetraphenylmorpholinochlorins, with and without direct ß-carbon-to-o-phenyl linkages to the flanking phenyl groups, is described. The morpholinochlorins, a class of stable chlorin analogues, were synthesized in two to three steps from meso-tetraphenylporphyrin. The conformations and the relative stereostructures of a variety of free-base and Ni(II) complexes of these morpholinochlorins were elucidated by X-ray diffractometry. Steric and stereoelectronic arguments explain the relative stereoarray of the morpholino-substituents, which differ in the free-base and Ni(II) complexes, and in the monoalkoxy, ß-carbon-to-o-phenyl linked morpholinochlorins, and the dialkoxy derivatives. The Ni(II) complexes were all found to be severely ruffled whereas the free-base chromophores are more planar. As a result of the helimeric distortion of their porphyrinoid chromophores, the ruffled macrocycles possess a stable inherent element of chirality. Most significantly, resolution of the racemic mixtures was achieved, both by classical methods via diastereomers and by HPLC on a chiral phase. Full CD spectra were recorded and modeled using quantum-chemical computational methods, permitting, for the first time, an assignment of the absolute configurations of the chromophores. The report expands the range of known pyrrole-modified porphyrins. Beyond this, it introduces large chiral porphyrinoid π-systems that exist in the form of two enantiomeric, stereochemically stable helimers that can be resolved. This forms the basis for possible future applications, for example, in molecular-recognition systems or in materials with chiroptic properties.

A role for the unfolded protein response (UPR) in virulence and antifungal susceptibility in Aspergillus fumigatus.

Filamentous fungi rely heavily on the secretory pathway, both for the delivery of cell wall components to the hyphal tip and the production and secretion of extracellular hydrolytic enzymes needed to support growth on polymeric substrates. Increased demand on the secretory system exerts stress on the endoplasmic reticulum (ER), which is countered by the activation of a coordinated stress response pathway termed the unfolded protein response (UPR). To determine the contribution of the UPR to the growth and virulence of the filamentous fungal pathogen Aspergillus fumigatus, we disrupted the hacA gene, encoding the major transcriptional regulator of the UPR. The DeltahacA mutant was unable to activate the UPR in response to ER stress and was hypersensitive to agents that disrupt ER homeostasis or the cell wall. Failure to induce the UPR did not affect radial growth on rich medium at 37 degrees C, but cell wall integrity was disrupted at 45 degrees C, resulting in a dramatic loss in viability. The DeltahacA mutant displayed a reduced capacity for protease secretion and was growth-impaired when challenged to assimilate nutrients from complex substrates. In addition, the DeltahacA mutant exhibited increased susceptibility to current antifungal agents that disrupt the membrane or cell wall and had attenuated virulence in multiple mouse models of invasive aspergillosis. These results demonstrate the importance of ER homeostasis to the growth and virulence of A. fumigatus and suggest that targeting the UPR, either alone or in combination with other antifungal drugs, would be an effective antifungal strategy.

Pioglitazone suppresses inflammation in vivo in murine carotid atherosclerosis: novel detection by dual-target fluorescence molecular imaging.

OBJECTIVE: To investigate the effects of pioglitazone (PIO), a peroxisome proliferator-activated receptor γ agonist, on plaque matrix metalloproteinase (MMP) and macrophage (Mac) responses in vivo in a molecular imaging study. METHODS AND RESULTS: In vitro, PIO suppressed MMP-9 protein expression in murine peritoneal Macs (P<0.05). To assess PIO's effects on plaque inflammation, nondiabetic apolipoprotein E(-/-) mice receiving a high-cholesterol diet (HCD) were administered an MMP-activatable fluorescence imaging agent and a spectrally distinct Mac-avid fluorescent nanoparticle. After 24 hours, mice underwent survival dual-target intravital fluorescence microscopy of carotid arterial plaques. These mice were then randomized to HCD or HCD plus 0.012% PIO for 8 weeks, followed by a second intravital fluorescence microscopy study of the same carotid plaque. In the HCD group, in vivo MMP and Mac target-to-background ratios increased similarly (P<0.01 versus baseline). In contrast, PIO reduced MMP and Mac target-to-background ratios (P<0.01) versus HCD. Changes in MMP and Mac signals correlated strongly (r ≥0.75). Microscopy demonstrated MMP and Mac reductions in PIO-treated mice and a PIO-modulated increase in plaque collagen. CONCLUSIONS: Serial optical molecular imaging demonstrates that plaque MMP and Mac activity in vivo intensify with hypercholesterolemia and are reduced by PIO therapy.

Bio-inspired nanomaterials as novel options for the treatment of cardiovascular disease.

Cardiovascular disease (CVD) and its sequelae have long been the leading causes of death and disability in the developed world. Although mortality associated with CVD has been decreasing, due in large part to novel therapeutic options, the rate of decrease has flattened. Thus, there is a great need to investigate alternate therapeutic strategies that can increase efficacy while decreasing adverse effects. Nanomaterials have been widely investigated and have emerged as promising tools for both therapeutic and diagnostic purposes in oncology; however, the potential of nanomaterials has not been extensively explored for cardiovascular medicine. In this review, we focus on recent developments in the field of nanomedicines targeted for CVDs, with a special emphasis on cell membrane-coated nanoparticles (NPs) and their applications.