Decrease In Lesion Size Research Articles

Innovative thrombolytic strategy using a heterodimer diabody against TAFI and PAI-1 in mouse models of thrombosis and stroke

AbstractCirculating thrombin-activatable fibrinolysis inhibitor (TAFI) and plasminogen activator inhibitor-1 (PAI-1) are causal factors for thrombolytic failure. Therefore, we evaluated an antibody-engineered bispecific inhibitor against TAFI and PAI-1 (heterodimer diabody, Db-TCK26D6x33H1F7) in several mouse models of thrombosis and stroke. Prophylactic administration of the diabody (0.8 mg/kg) in a thromboplastin-induced model of thromboembolism led to decreased lung fibrin deposition. In a model of cerebral ischemia and reperfusion, diabody administration (0.8 mg/kg, 1 hour postocclusion) led to a mitigated cerebral injury with a 2.3-fold reduced lesion and improved functional outcomes. In a mouse model of thrombin-induced middle cerebral artery occlusion, the efficacy of the diabody was compared to the standard thrombolytic treatment with recombinant tissue-type plasminogen activator (tPA). Early administration of diabody (0.8 mg/kg) caused a twofold decrease in brain lesion size, whereas that of tPA (10 mg/kg) had a much smaller effect. Delayed administration of diabody or tPA had no effect on lesion size, whereas the combined administration of diabody with tPA caused a 1.7-fold decrease in lesion size. In contrast to tPA, the diabody did not increase accumulative bleeding. In conclusion, administration of a bispecific inhibitor against TAFI and PAI-1 results in a prominent profibrinolytic effect in mice without increased bleeding.

Abstract T P79: Rapamycin Improves Outcome Post Stroke Injury by Enhancing Autophagy in Neuronal Cells

Background and Purpose: Previous literature and studies from our laboratory indicate that pharmacological inhibition and enhancement of autophagy are both effective in reducing infarct volume after stroke injury. Methods: Mice were treated with rapamycin to enhance, and chloroquine to suppress autophagy in mice immediately after stroke injury and again 24 hours post stroke injury in a murine stroke model. Survival data and neurological score were taken before sacrifice at 48 hours and lesion size was determined by TTC. HT22 neuronal mouse hippocampal cells were treated for one hour with H2O2 to mimic an oxidative stress injury and treated concurrently with rapamycin to enhance autophagy, chloroquine to inhibit autophagy or ZVAD to inhibit apoptosis. The treatment with pharmacological agents continued for 24 hours. Cells were counted with trypan blue on a hemocytometer. Western blot analysis was used to monitor protein changes. shRNA to Atg5 was used to genetically inhibit autophagy. Results: Both pharmacological agents showed decreased lesion size, though enhancement of autophagy with rapamycin exhibited a greater reduction in lesion size as well as increased survival and an improved neurological score. Rapamycin improved survival of neurons after oxidative injury and genetic inhibition of autophagy abrogated rapamycin's positive effect. Conclusions: Rapamycin improves stroke outcome by enhancing autophagy in neuronal cells.

Acute Disseminated Encephalomyelitis Responsive to Cyclophosphamide Therapy: A Case Report

Acute disseminated encephalomyelitis (ADEM) is a rare neurologic condition that affects the central nervous system (CNS). In this report, a 22-year-old fraternal twin male developed ADEM 6 months post viral infection. He presented non-verbal with doll’s eyes, a right facial droop, right-sided upper and lower extremity weakness, as well as brisk reflexes and clonus on the right. He failed to improve with traditional therapies such as intravenous steroids, intravenous immunoglobulin, and plasmapheresis. However, 2 days following a 3-day course of intravenous cyclophosphamide 1,000 mg/m 2 , he spontaneously opened his eyes, developed masticatory movements, mouthed words, and regained movement in his extremities against gravity. There was a 40.7% and 16.6% decrease in lesion area in brain magnetic resonance imaging (MRI) T2 FLAIR sagittal and axial images, respectively over that 2-day period. This is the first reported case that shows neurological improvement, measured by the functional independence measure (FIM) scale, in addition to decreased lesion size on brain MRI T2 FLAIR images after implementation of cyclophosphamide therapy. Cyclophosphamide should be considered as an alternative therapy in patients with refractory ADEM. J Med Cases. 2015;6(7):290-294 doi: https://doi.org/10.14740/jmc2176w

Effect of pharmacologic resuscitation on the brain gene expression profiles in a swine model of traumatic brain injury and hemorrhage.

We have previously shown that addition of valproic acid (VPA; a histone deacetylase inhibitor) to hetastarch (Hextend [HEX]) resuscitation significantly decreases lesion size in a swine model of traumatic brain injury (TBI) and hemorrhagic shock (HS). However, the precise mechanisms have not been well defined. As VPA is a transcriptional modulator, the aim of this study was to investigate its effect on brain gene expression profiles. Swine were subjected to controlled TBI and HS (40% blood volume), kept in shock for 2 hours, and resuscitated with HEX or HEX + VPA (n = 5 per group). Following 6 hours of observation, brain RNA was isolated, and gene expression profiles were measured using a Porcine Gene ST 1.1 microarray (Affymetrix, Santa Clara, CA). Pathway analysis was done using network analysis tools Gene Ontology, Ingenuity Pathway Analysis, and Parametric Gene Set Enrichment Analysis. Real-time polymerase chain reaction was used to verify the key microarray findings. A total of 1,668 probe sets mapping to 370 known genes were differentially expressed between the HEX and HEX + VPA groups. Expression of apoptotic genes differed between groups, and biologic function analysis predicted a significant downregulation of apoptosis (p = 1.29 × 10), cell death (p = 8.46 × 10), and necrosis (p = 9.07 × 10). Pathway analysis indicated a significant modulation of pathways involved in cell signaling, dendritic cell response, and the complement system. This is the first high-throughput analysis of cerebral gene profiling following TBI + HS. It shows that treatment with VPA significantly alters early transcription of pathways related to cell survival, which may explain its neuroprotective effects.

Phospholipid transfer protein destabilizes mouse atherosclerotic plaque.

Phospholipid transfer protein (PLTP) accelerates the development of atherosclerosis in mouse models. We examined the role of PLTP in atherosclerotic plaque stability. We prepared apolipoprotein E and PLTP double-knockout (PLTP(-/-)ApoE(-/-)) mice. PLTP deficiency significantly decreased lesion size and reduced monocyte/macrophage infiltration, as well as macrophage apoptosis in lesion areas. Moreover, it increased fibrous content in plaques, which suggests that PLTP may affect atherosclerotic plaque stability. Importantly, PLTP overexpression mediated by adenovirus had the reverse effect. It promoted the accumulation of reactive oxygen species in macrophages, which could lead to cell apoptosis and increased the production of inflammatory cytokines and chemokines. PLTP overexpression could promote receptor-interacting protein 3 recruitment of macrophages in cytoplasm, which could induce reactive oxygen species, thus inducing atherogenesis. PLTP plays an important role in modulating the stability of atherosclerotic plaques. The receptor-interacting protein 3- reactive oxygen species signal pathway could be involved in this PLTP-mediated process.

Ouabain improves functional recovery following traumatic brain injury.

The cardiac steroid ouabain binds to Na(+), K(+)-ATPase and inhibits its activity. Administration of the compound to animals and humans causes an increase in the force of contraction of heart muscle and stabilizes heart rate. In addition, this steroid promotes the growth of cardiac, vascular, and neuronal cells both in vitro and in vivo. We studied the effects of ouabain on mouse recovery following closed head injury (CHI), a model for traumatic brain injury. We show that chronic (three times a week), but not acute, intraperitoneal administration of a low dose (1 μg/kg) of ouabain significantly improves mouse recovery and functional outcome. The improvement in mouse performance was accompanied by a decrease in lesion size, estimated 43 d following the trauma. In addition, mice that underwent CHI and were treated with ouabain showed an increase in the number of proliferating cells in the subventricular zone and in the area surrounding the site of injury. Determination of the identity of the proliferating cells in the area surrounding the trauma showed that whereas there was no change in the proliferation of endothelial cells or astrocytes, neuronal cell proliferation almost doubled in the ouabain-treated mice in comparison with that of the vehicle animals. These results point to a neuroprotective effects of low doses of ouabain and imply its involvement in brain recovery and neuronal regeneration. This suggests that ouabain and maybe other cardiac steroids may be used for the treatment of traumatic brain injury.

PO-0472 Evaluation Of Secretoneurin As Therapeutic Strategy In Neonatal Excitotoxic Brain Injury

Background Preterm brain injury causes neurodevelopmental disability. Excitotoxicity plays an important role in the pathogenesis of preterm brain injury. The neuropeptide secretoneurin (SN) was shown to enhance angiogenesis and neurogenesis in a rodent model of adult ischaemic brain damage. SN might be considered a promising substance in newborn brain injury. Aim To evaluate the effect of SN as a therapeutic strategy in an established in vivo model of excitotoxic newborn brain injury. Methods Five-day-old mice pups were subjected to an intracranial injection of ibotenic acid into the right brain hemisphere. After recovery for one hour, animals were randomly treated with an intraperitoneal injection of i) vehicle, ii) SN 0.25 µg/g body weight (bw) or iii) SN 2.5 µg/g bw. Brains were harvested 24 and 120 h after the insult and processed for histological analysis. As a primary outcome parameter lesion size in cortical grey and white matter was evaluated. Results SN administration had no significant effect on lesion size 120 h after the insult. When evaluated 24 h after the excitotoxic insult, SN showed a marked, but non-significant trend towards a decreased lesion size in white matter (SN 0.25 µg/g 323.33 ± 128.82, SN 2.5 µg/g bw 298.46 ± 137.46, vehicle 440.00 ± 227.81, n = 13–19, p = 0.06). SN had no effect on lesion size in grey matter. Conclusion This study shows a trend towards a reduced injury in white matter in an in vivo model of neonatal brain injury. We are currently performing immunohistochemical analysis to evaluate underlying mechanisms that could result in long-term protection.

Mesenchymal stem cells overexpressing IL-13 decrease lesion size and demyelination after spinal cord injury

[Dooley, Dearbhaile; Lemmens, Evi; Vangansewinkel, Tim; Lemmens, Stefanie; Hendrix, Sven] Hasselt Univ, Biomed Res Inst, Diepenbeek, Belgium. [De Vocht, Nathalie; Le Blon, Debbie; Ponsaerts, Peter] Univ Antwerp, Lab Expt Hematol, Vaccine & Infect Dis Inst Vaxinfectio, B-2020 Antwerp, Belgium.

Management of Central Giant Cell Granuloma With Subcutaneous Denosumab Therapy

In recent years, the treatment of central giant cell granuloma (CGCG) has become focused on the inhibition of osteoclast differentiation and proliferation. Medications that were developed for the treatment of giant cell tumor of bone and bone resorption from metastatic skeletal disease have shown some success in the treatment of CGCG. The present report describes 2 cases of CGCG of the mandible that were treated effectively with subcutaneous denosumab. Two cases of histologically diagnosed CGCG of the mandible were treated with monthly subcutaneous injections of denosumab 120 mg primarily or after intralesional corticosteroid therapy. Clinical and radiographic follow-ups were recorded over a period of 24 months (case 1) and 15months (case 2). In the 2 cases, progressive radiodensity and osseous regeneration were noted 4 to 6 months after denosumab therapy was initiated. A decrease in lesion size and improvement in bone contour and facial symmetry were seen in the 2 cases. The major radiographic, clinical, and histologic responses seen in these 2 cases suggest that denosumab may represent a viable alternative or adjunctive procedure to eliminate or decrease the extent of surgical intervention and morbidity in the treatment of CGCG. Future prospective studies with a larger sample would provide more comprehensive information about the long-term effects and possible adverse side effects of treating CGCG of the jaws with denosumab.

Rapamycin up-regulation of autophagy reduces infarct size and improves outcomes in both permanent MCAL, and embolic MCAO, murine models of stroke

Background and purposeThe role of autophagy in response to ischemic stroke has been confusing with reports that both enhancement and inhibition of autophagy decrease infarct size and improve post-stroke outcomes. We sought to clarify this by comparing pharmacologic modulation of autophagy in two clinically relevant murine models of stroke.MethodsWe used rapamycin to induce autophagy, and chloroquine to block completion of autophagy, by treating mice immediately after stroke and at 24 hours post-stroke in two different models; permanent Middle Cerebral Artery Ligation (MCAL), which does not allow for reperfusion of distal trunk of middle cerebral artery, and Embolic Clot Middle Cerebral Artery Occlusion (eMCAO) which allows for a slow reperfusion similar to that seen in most human stroke patients. Outcome measures at 48 hours post-stroke included infarct size analysis, behavioral assessment using Bederson neurological scoring, and survival.ResultsChloroquine treatment reduced the lesion size by approximately 30% and was significant only in the eMCAO model, where it also improved the neurological score, but did not increase survival. Rapamycin reduced lesion size by 44% and 50% in the MCAL and eMCAO models, respectively. Rapamycin also improved the neurological score to a greater degree than chloroquine and improved survival.ConclusionsWhile both inhibition and enhancement of autophagy by pharmacological intervention decreased lesion size and improved neurological scores, the enhancement with rapamycin showed a greater degree of improvement in outcomes as well as in survival. The protective action seen with chloroquine may be in part due to off-target effects on apoptosis separate from blocking lysosomal activity in autophagy. We conclude pharmacologic induction of autophagy is more advantageous than its blockade in physiologically-relevant permanent and slow reperfusion stroke models.

Antileishmanial activity of the estrogen receptor modulator raloxifene.

BackgroundThe treatment of leishmaniasis relies mostly on parenteral drugs with potentially serious adverse effects. Additionally, parasite resistance in the treatment of leishmaniasis has been demonstrated for the majority of drugs available, making the search for more effective and less toxic drugs and treatment regimens a priority for the control of leishmaniasis. The aims of this study were to evaluate the antileishmanial activity of raloxifene in vitro and in vivo and to investigate its mechanism of action against Leishmania amazonensis.Methodology/Principal FindingsRaloxifene was shown to possess antileishmanial activity in vitro against several species with EC50 values ranging from 30.2 to 38.0 µM against promastigotes and from 8.8 to 16.2 µM against intracellular amastigotes. Raloxifene's mechanism of action was investigated through transmission electron microscopy and labeling with propidium iodide, DiSBAC2(3), rhodamine 123 and monodansylcadaverine. Microscopic examinations showed that raloxifene treated parasites displayed autophagosomes and mitochondrial damage while the plasma membrane remained continuous. Nonetheless, plasma membrane potential was rapidly altered upon raloxifene treatment with initial hyperpolarization followed by depolarization. Loss of mitochondrial membrane potential was also verified. Treatment of L. amazonensis – infected BALB/c mice with raloxifene led to significant decrease in lesion size and parasite burden.Conclusions/SignificanceThe results of this work extend the investigation of selective estrogen receptor modulators as potential candidates for leishmaniasis treatment. The antileishmanial activity of raloxifene was demonstrated in vitro and in vivo. Raloxifene produces functional disorder on the plasma membrane of L. amazonensis promastigotes and leads to functional and morphological disruption of mitochondria, which culminate in cell death.

Investigating the Clinical Potential for 14-3-3 Zeta Protein to Serve as a Biomarker for Epithelioid

INTRODUCTION: Recently, 14-3-3 zeta protein was identified as a potential serum biomarker of epithelioid ovarian cancer. The goal of this study was to investigate the clinical potential of 14-3-3 zeta protein for monitoring epithelioid ovarian cancer progression compared with CA-125 and HE4. In a prospective follow-up study, carried out at the University of Pecs Medical Center Department of Obstetrics and Gynecology/Oncology (Pecs, Hungary), we investigated 13 patients with epithelioid ovarian cancer with advanced-stage (International Federation of Gynecology and Obstetrics IIb–IIIc) epitheloid ovarian cancer who underwent radical surgery and received six consecutive cycles of first-line chemotherapy (paclitaxel, carboplatin) in 21-day intervals. METHODS: Pre- and postchemotherapy computed tomography scans were performed. Serum levels of CA-125, HE4, and 14-3-3 zeta protein were detected by enzyme-linked immunosorbent assay and quantitative electrochemiluminescence assay. RESULTS: Serum levels of CA-125, HE4, and 14-3-3 zeta protein as well as lesion size according to pre- and postchemotherapy computed tomography scans were analyzed. Serum levels of CA-125 and HE4 were found to significantly decrease after chemotherapy, and this was consistent with the decrease in lesion size detected postchemotherapy. In contrast, 14-3-3 zeta protein levels did not significantly differ in healthy postmenopausal patients compared with patients with epithelioid ovarian cancer. CONCLUSIONS: Determination of CA-125 and HE4 serum levels for the risk of ovarian malignancy algorithm does represent a useful tool for the prediction of chemotherapy efficacy for patients with epithelioid ovarian cancer. However, levels of 14-3-3 zeta protein were not found to be a reliable biomarker of epithelioid ovarian cancer.

Abstract 246: Anacetrapib Adds to the Antiatherogenic Effect of Atorvastatin, Mainly by Increasing the Clearance of Non-HDL Cholesterol

Introduction: The residual risk that remains after statin treatment has stimulated the search for secondary treatment targets. Epidemiological studies propose HDL-C as a possible candidate. Cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from atheroprotective HDL to atherogenic (V)LDL. The CETP inhibitor anacetrapib decreases (V)LDL-C by ~15-40% and increases HDL-C by ~40-140% in clinical trials. We evaluated the effects of anacetrapib on lipid metabolism, HDL function and atherosclerosis, and examined possible additive/synergistic effects of anacetrapib on top of atorvastatin in APOE*3Leiden.CETP mice. Methods and results: Mice were fed a diet without or with ascending dosages of anacetrapib (0.03; 0.3; 3; 30 mg/kg/d), atorvastatin (2.4 mg/kg/d) alone or in combination with anacetrapib (0.3 mg/kg/d) for 21 weeks. Anacetrapib dose-dependently reduced CETP activity (-59% to -100%, P<0.01), thereby decreasing nonHDL-C (-24% to -45%, P<0.001) and increasing HDL-C (+30% to +86%, P<0.001). Anacetrapib did not affect HDL function. Additionally, we showed that anacetrapib reduced nonHDL-C mainly by increasing the clearance by the liver. Anacetrapib dose-dependently reduced atherosclerotic lesion area (-41% to -92%, P<0.01) and severity, increased the plaque stability index and added to the effects of atorvastatin by further decreasing lesion size (-95%, P<0.001) and severity. Analysis of covariance showed that both anacetrapib (P<0.05) and nonHDL-C (P<0.001), but not HDL-C (P=0.76), independently decreased lesion size. Conclusions: Anacetrapib reduces atherosclerosis, and adds to the anti-atherogenic effect of atorvastatin. The anti-atherogenic effect is mainly ascribed to a reduction in nonHDL-C, as a consequence of an increased clearance. In addition, anacetrapib improves lesion stability.

Abstract 676: 14q32 MicroRNA Inhibition Reduces Atherosclerotic Lesion Formation and Increases Plaque Stability

Introduction: Atherosclerosis is a multifactorial disease involving inflammatory, metabolic and lipid-related processes. As microRNAs (miRs) regulate expression of up to hundreds of target genes, we aimed to identify miRs that target genes involved in all processes of atherosclerosis. Using www.targetscan.org, we performed a Reverse Target Prediction on a set of 150 atherosclerosis-related genes. We found enrichment of binding sites for several miRs from a single gene cluster on human chromosome 14. Methods: ApoE-/- mice fed a Western type diet received semi-constrictive collars around both carotid arteries to induce atherosclerotic lesion formation. Gene Silencing Oligonucleotides (GSOs) were used to inhibit 3 14q32 miRs: miR-329, miR-494 and miR-495. Mice were injected with 1 mg GSO at day 4 after collar placement and again with 0.5 mg GSO at day 18. Mice were sacrificed at day 7 and day 28 for analysis of carotid arteries, liver, spleen, bone marrow and blood samples. Results: Atherosclerotic plaque analysis revealed a striking decrease of lesion size in mice treated with GSO-494 and GSO-495 (GSO-Control: 47 ± 11*103 μm2; GSO-494: 16 ± 3*103 μm2; GSO-495: 22 ± 5*103 μm2), whereas a trend towards reduced plaque size was observed in mice treated with GSO-329 (27 ± 6*103 μm2). Moreover, plaque stability was increased in mice treated with GSO-494 and GSO-495, determined by both a decrease in necrotic core size (GSO-494: 80% decrease; GSO-495: 60% decrease compared to GSO-Control) and an increase in plaque collagen content (GSO-Control: 6.6 ± 1.6%; GSO-494: 12.7 ± 2.1%; GSO-495: 15.8 ± 3.1%). Furthermore, inhibition of miR-494 and miR-495 resulted in decreased plasma cholesterol levels (GSO-Control: 30.4 ± 1.1 mM; GSO-494: 26.4 ± 0.7 mM; GSO-495: 26.4 ± 1.6) and decreased VLDL fractions. Conclusions: Inhibition of miR-494, miR-495 and, to a lesser extent, miR-329, leads to reduced plaque size, increased plaque stability and reduced plasma cholesterol levels. This makes 14q32 miRs promising novel therapeutic targets in atherosclerosis.

Abstract 653: Endothelial ASC Modulates Early Atherosclerosis by Limiting Monocyte Recruitment

The inflammasome consists of an innate immune receptor, the adaptor molecule ASC, and the effector Caspase-1 that is responsible for processing signature cytokines such as IL-1beta. The exact role of the inflammasome in atherosclerosis is not clear since both pathogenic and dispensable roles have been reported. Notably, ASC has functions that may impact atherosclerosis but are unrelated to the inflammasome, for example the induction of apoptosis and the regulation of pro-inflammatory transcription factors. The various roles of ASC (i.e. IL-1beta processing, cell death, NF-kappaB and AP-1 regulation) may modulate the progression of atherosclerosis depending on lesion stage and explain the divergent findings highlighted above. To examine the role of ASC in lesion progression, we bred ASC -deficient mice onto the Ldlr -/- background and initiated a cholesterol-rich diet (CRD). In the early stages of atherogenesis (2 weeks of CRD), ASC -/- Ldlr -/- mice had a significantly increased lesion surface area, leukocyte content, and rate of Ly6C hi monocyte recruitment in the ascending aorta. Bone marrow transplant experiments suggested that ASC deficiency in the vessel wall was responsible for this phenotype. Knockdown of ASC by siRNA in cultured human umbilical vein endothelial cells lead to increased TNF-alpha-induced expression of VCAM1 and monocyte adhesion in parallel-plate flow chamber assays. Monocyte adhesion was not affected by silencing Caspase-1, suggesting that the effect of ASC was independent of the inflammasome. Preliminary findings implicate MAPK family members in ASC-mediated regulation of endothelial adhesion molecules. At 16 weeks of a CRD ASC -/- Ldlr -/- mice had a significantly decreased lesion size compared with controls. Lesions from ASC -/- Ldlr -/- mice had a smaller necrotic core area accompanied by decreased numbers of apoptotic leukocytes as determined by active Caspase-3 and TUNEL positivity. These data are in accordance with the pro-atherogenic role of ASC in later stages of atherosclerosis. However our findings demonstrate a novel protective role for ASC in the early stage, which represents a non-inflammasome function via modulation of endothelial-associated adhesion molecules.

Abstract 655: 4-Phenylbutyrate in Drinking Water Reduced Atherosclerotic Lesion Growth in ApoE-/- Mice by Modifying Monocyte-Macrophage Differentiation

Cardiovascular disease and atherothrombosis account for the majority of deaths worldwide. Plaque rupture, thrombus formation, and vessel occlusion contribute to the acute clinical manifestations of atherothrombosis. Previously we have shown that the unfolded protein response (UPR) has an important role in monocyte-macrophage differentiation. Furthermore, macrophage infiltration, lipid accumulation and foam cell formation are key events in atherogenesis. Endoplasmic reticulum (ER) stress-dependent apoptosis also contributes to atherosclerotic lesion growth and plaque necrosis. The aim of the current study was to examine whether 4-phenylbutyrate (4-PBA), a small chemical chaperone that alleviates ER stress, confers protection against atherosclerosis when provided via drinking water. To investigate the ability of 4-PBA to affect atherosclerotic lesion growth, at 12 weeks of age female ApoE-/- mice on chow diet were provided with 4-PBA (1 - 2.5 g/kg/d) in their drinking water for five weeks, ad libitum. Following treatment, mice were sacrificed, their tissues collected and atherosclerotic lesions examined by immunohistochemistry. Early atherosclerotic lesion area was significantly reduced in ApoE-/- mice provided 4-PBA-treated water from 12 - 17 weeks of age, relative to controls (0.6 x105 ± 0.1 μm2 versus 1.1 x105 ± 0.1 μm2, respectively, P < 0.05). We hypothesized that 4-PBA drinking water decreased lesion size by a mechanism involving reduced ER stress and monocyte-macrophage differentiation. In support of this mechanism, preliminary data indicates 4-PBA inhibited monocyte-macrophage differentiation, as measured by mRNA expression of the macrophage marker MSR-1 (0.5 ± 0.2 fold-change vs. control) and cell adherence (37 ± 5% vs. control) in PMA-treated THP-1 cells. In summary, these findings provide evidence that 4-PBA-treated drinking water reduced atherosclerotic lesion size in a mouse model of atherosclerosis. We hypothesize that 4-PBA acts in part through its ability to reduce ER stress and modulate monocyte/macrophage differentiation and function. Additional studies are in progress to further examine the mechanisms by which 4-PBA affects atherogenesis.

Multimodal Imaging for Early Functional Response Assessment of 90Y-/177Lu-DOTATOC Peptide Receptor Targeted Radiotherapy with DW-MRI and 68Ga-DOTATOC-PET/CT

The purpose of this study is to investigate the utility of contrast-enhanced magnetic resonance imaging (CE-MRI), diffusion-weighted MRI (DW-MRI), and (68)Ga-DOTATOC positron emission tomography/computer tomography ((68)Ga-DOTATOC PET/CT) in the assessment of response to loco-regional peptide receptor radiotherapy (PRRT) with (90)Y-/(177)Lu-DOTATOC in patients with hepatic metastases from gastro-entero-pancreatic neuroendocrine tumors (GEP-NET). CE-MRI, DW-MRI, and (68)Ga-DOTATOC-PET/CT images were acquired before and 3months after one to two cycles of intra-arterial (90)Y-/(177)Lu-DOTATOC therapy in 14 patients (nine female, five male; mean age, 54 ± 9years; range, 41-69years) with hepatic metastases from GEP-NET. A total of 38 liver metastases were defined as target lesions for which the longest diameter, mean apparent diffusion coefficient (ADCmean) and maximum standardized uptake value (SUVmax) were assessed. Based on changes in size on follow-up imaging, target lesions were classified as responding (RL) or nonresponding (NRL). Relative changes in tumor size, ADCmean, and SUVmax were compared between the two subgroups. A total of 27 responding and 11 nonresponding lesions were successfully evaluated. Mean ADCmean increased significantly in RL (p = 0.011) as well as NRL (p = 0.025). A significant correlation was found between baseline ADCmean and both the percent ADCmean change (p = 0.033) and decrease in lesion size after therapy (diameter p = 0.006; volume p = 0.002). SUVmax of RL declined significantly by 24.1% (p = 0.014) and remained nearly unchanged in NRL. The change of SUVmax correlated significantly with the pretreatment SUVmax (p < 0.001) and the change in lesion diameter (p = 0.009). NRL with an ADCmean change >0.31 × 10(-3)mm(2)/s on first follow-up imaging showed a decrease in size in the long-term course. These results suggest that both DW-MRI and DOTATOC-PET imaging provide potential biomarkers for early assessment of treatment and stratification of therapy response, but that DW-MRI should be interpreted only in combination with SSTR expression and morphologic changes.

Comparison of the Effect of Minocycline and Simvastatin on Functional Recovery and Gene Expression in a Rat Traumatic Brain Injury Model

The goal of this study was to compare the effects of minocycline and simvastatin on functional recovery and brain gene expression after a cortical contusion impact (CCI) injury. Dosage regimens were designed to provide serum concentrations in a rat model in the range obtained with clinically approved doses; minocycline 60 mg/kg q12h and simvastatin 10 mg/kg q12h for 72 h. Functional recovery was assessed using motor and spatial learning tasks and neuropathological measurements. Microarray-based transcriptional profiling was used to determine the effect on gene expression at 24 h, 72 h, and 7 days post-CCI. Gene Ontology analysis (GOA) was used to evaluate the effect on relevant biological pathways. Both minocycline and simvastatin improved fine motor function, but not gross motor or cognitive function. Minocycline modestly decreased lesion size with no effect of simvastatin. At 24 h post-CCI, GOA identified a significant effect of minocycline on chemotaxis, blood circulation, immune response, and cell to cell signaling pathways. Inflammatory pathways were affected by minocycline only at the 72 h time point. There was a minimal effect of simvastatin on gene expression 24 h after injury, with increasing effects at 72 h and 7 days. GOA identified a significant effect of simvastatin on inflammatory response at 72 h and 7 days. In conclusion, treatment with minocycline and simvastatin resulted in significant effects on gene expression in the brain reflecting adequate brain penetration without producing significant neurorestorative effects.

TREM-1 Deficiency Can Attenuate Disease Severity without Affecting Pathogen Clearance

Triggering receptor expressed on myeloid cells-1 (TREM-1) is a potent amplifier of pro-inflammatory innate immune reactions. While TREM-1-amplified responses likely aid an improved detection and elimination of pathogens, excessive production of cytokines and oxygen radicals can also severely harm the host. Studies addressing the pathogenic role of TREM-1 during endotoxin-induced shock or microbial sepsis have so far mostly relied on the administration of TREM-1 fusion proteins or peptides representing part of the extracellular domain of TREM-1. However, binding of these agents to the yet unidentified TREM-1 ligand could also impact signaling through alternative receptors. More importantly, controversial results have been obtained regarding the requirement of TREM-1 for microbial control. To unambiguously investigate the role of TREM-1 in homeostasis and disease, we have generated mice deficient in Trem1. Trem1−/− mice are viable, fertile and show no altered hematopoietic compartment. In CD4+ T cell- and dextran sodium sulfate-induced models of colitis, Trem1−/− mice displayed significantly attenuated disease that was associated with reduced inflammatory infiltrates and diminished expression of pro-inflammatory cytokines. Trem1−/− mice also exhibited reduced neutrophilic infiltration and decreased lesion size upon infection with Leishmania major. Furthermore, reduced morbidity was observed for influenza virus-infected Trem1−/− mice. Importantly, while immune-associated pathologies were significantly reduced, Trem1−/− mice were equally capable of controlling infections with L. major, influenza virus, but also Legionella pneumophila as Trem1+/+ controls. Our results not only demonstrate an unanticipated pathogenic impact of TREM-1 during a viral and parasitic infection, but also indicate that therapeutic blocking of TREM-1 in distinct inflammatory disorders holds considerable promise by blunting excessive inflammation while preserving the capacity for microbial control.

Activation of Liver X Receptor Decreases Atherosclerosis in Ldlr −/− Mice in the Absence of ATP-Binding Cassette Transporters A1 and G1 in Myeloid Cells

Liver X receptor (LXR) activators decrease atherosclerosis in mice. LXR activators (1) directly upregulate genes involved in reverse cholesterol transport and (2) exert anti-inflammatory effects mediated by transrepression of nuclear factor-κB target genes. We investigated whether myeloid cell deficiency of ATP-binding cassette transporters A1 and G1 (ABCA1/G1), principal targets of LXR that promote macrophage cholesterol efflux and initiate reverse cholesterol transport, would abolish the beneficial effects of LXR activation on atherosclerosis. LXR activator T0901317 substantially reduced inflammatory gene expression in macrophages lacking ABCA1/G1. Ldlr(-/-) mice were transplanted with Abca1(-/-)Abcg1(-/-) or wild-type bone marrow (BM) and fed a Western-type diet for 6 weeks with or without T0901317 supplementation. Abca1/g1 BM deficiency increased atherosclerotic lesion complexity and inflammatory cell infiltration into the adventitia and myocardium. T0901317 markedly decreased lesion area, complexity, and inflammatory cell infiltration in the Abca1(-/-)Abcg1(-/-) BM-transplanted mice. To investigate whether this was because of macrophage Abca1/g1 deficiency, Ldlr(-/-) mice were transplanted with LysmCreAbca1(fl/fl)Abcg1(fl/fl) or Abca1(fl/fl)Abcg1(fl/fl) BM and fed Western-type diet with or without the more specific LXR agonist GW3965 for 12 weeks. GW3965 decreased lesion size in both groups, and the decrease was more prominent in the LysmCreAbca1(fl/fl)Abcg1(fl/fl) group. The results suggest that anti-inflammatory effects of LXR activators are of key importance to their antiatherosclerotic effects in vivo independent of cholesterol efflux pathways mediated by macrophage ABCA1/G1. This has implications for the development of LXR activators that lack adverse effects on lipogenic genes while maintaining the ability to transrepress inflammatory genes.