Oligodendrocytes are susceptible to Zika virus infection in a mouse model of perinatal exposure: Implications for CNS complications.

Some children with proven intrauterine Zika virus (ZIKV) infection who were born asymptomatic subsequently manifested neurodevelopmental delays, pointing to impairment of development perinatally and postnatally. To model this, we infected postnatal day (P) 5-6 (equivalent to the perinatal period in humans) susceptible mice with a mammalian cell-propagated ZIKV clinical isolate from the Brazilian outbreak in 2015. All infected mice appeared normal up to 4 days post-intraperitoneal inoculation (dpi), but rapidly developed severe clinical signs at 5-6 dpi. All nervous tissue examined at 5/6 dpi appeared grossly normal. However, anti-ZIKV positive cells were observed in the optic nerve, brain, and spinal cord; predominantly in white matter. Co-labeling with cell type specific markers demonstrated oligodendrocytes and astrocytes support productive infection. Rarely, ZIKV positive neurons were observed. In spinal cord white matter, which we examined in detail, apoptotic cells were evident; the density of oligodendrocytes was significantly reduced; and there was localized microglial reactivity including expression of the NLRP3 inflammasome. Together, our observations demonstrate that a clinically relevant ZIKV isolate can directly impact oligodendrocytes. As primary oligodendrocyte cell death can lead later to secondary autoimmune demyelination, our observations may help explain neurodevelopmental delays in infants appearing asymptomatic at birth and commend lifetime surveillance.

In vitro evidence consistent with an interaction between wild-type and mutant SOD1 protein associated with canine degenerative myelopathy.

Canine degenerative myelopathy (DM) is a progressive neurological disorder that may be considered to be a large animal model for specific forms of the fatal human disease, familial amyotrophic lateral sclerosis (fALS). DM is associated with a c118G>A mutation of the superoxide dismutase 1 (Sod1) gene, and a significant proportion of cases are inherited in an autosomal recessive manner in contrast to the largely, but not exclusively, dominant mode of inheritance in fALS. The consensus view is that these Sod1/SOD1 mutations result in a toxic gain of function but the mechanisms remain unclear. Here we used an in vitro neuroblastoma cell line transfection system to monitor wild-type and mutant forms of SOD1 fusion proteins containing either a Cherry or an enhanced green fluorescent protein (EGFP) tag. These fusion proteins retained SOD1 enzymatic activity on a native gel assay system. We demonstrate that SOD1 aggregate density is significantly higher in DM transfectants compared to wild-type. In addition, we show by co-immunoprecipitation and confocal microscopy, evidence for a potential interaction between wild-type and mutant forms of SOD1 in co-transfected cells. While in vitro studies have shown SOD1 heterodimer formation in fALS models, this is the first report for DM SOD1. Therefore, despite for the majority of cases there is a difference in the mode of inheritance between fALS and DM, a similar interaction between wild-type and mutant SOD1 forms can occur. Clarifying the role of SOD1 in DM may also be of benefit to understanding the role of SOD1 in fALS.

Developing a Comprehensive Animal Care Occupational Health and Safety Program at a Land-Grant Institution.

The Public Health Service Policy on the Humane Care and Use of Laboratory Animals and sound ethical practices require institutions to provide safe working environments for personnel working with animals; this mandate is achieved in part by establishing an effective animal care Occupational Health and Safety Program (OHSP). Land-grant institutions often face unique organizational challenges in fulfilling this requirement. For example, responsibilities for providing health and safety programs often have historically been dispersed among many different divisions scattered around the campus. Here we describe how our institutional management personnel overcame organizational structure and cultural obstacles during the formation of a comprehensive campus-wide animal care OHSP. Steps toward establishing the animal care OHSP included assigning overall responsibility, identifying all stakeholders, creating a leadership group, and hiring a fulltime Animal Care OHSP Specialist. A web-based portal was developed, implemented, and refined over the past 7 y and reflected the unique organizational structures of the university and the needs of our research community. Through this web-based portal, hazards are identified, risks are assessed, and training is provided. The animal care OHSP now provides easy mandatory enrollment, supports timely feedback regarding hazards, and affords enrollees the opportunity to participate in voluntary medical surveillance. The future direction and development of the animal care OHSP will be based on the research trends of campus, identification of emerging health and safety hazards, and ongoing evaluation and refinement of the program.

Development of learning objectives for neurology in a veterinary curriculum: part II: postgraduates.

BACKGROUND: Specialization in veterinary medicine in Europe is organized through the Colleges of the European Board of Veterinary Specialization. To inform updating of the curriculum for residents of the European College of Veterinary Neurology (ECVN) job analysis was used. Defining job competencies of diploma holders in veterinary neurology can be used as references for curriculum design of resident training. With the support of the diplomates of the ECVN and the members of the European Society of Veterinary Neurology (ESVN) a mixed-method research, including a qualitative search of objectives and quantitative ranking with 149 Likert scale questions and 48 free text questions in 9 categories in a survey was conducted. In addition, opinions of different groups were subjected to statistical analysis and the result compared. RESULTS: A return rate of 62% (n = 213/341) was achieved. Of the competencies identified by the Delphi process, 75% objectives were expected to attain expert level; 24% attain advanced level; 1% entry level. In addition, the exercise described the 11 highly ranked competencies, the 3 most frequently seen diseases of the central and peripheral nervous systems and the most frequently used immunosuppressive, antiepileptic and chemotherapeutic drugs. CONCLUSION: The outcomes of this "Delphi job analysis" provide a powerful tool to align the curriculum for ECVN resident training and can be adapted to the required job competencies, based on expectations. The expectation is that for majority of these competencies diplomates should attain an expert level. Besides knowledge and clinical skills, residents and diplomates are expected to demonstrate high standards in teaching and communication. The results of this study will help to create a European curriculum for postgraduate education in veterinary neurology.

Resident experience increases diagnostic rate of thyroid fine-needle aspiration biopsies.

RATIONALE AND OBJECTIVES: The aim of this study was to determine whether the diagnostic yield of thyroid fine-needle aspirations (FNAs) changes over the course of residency training. MATERIALS AND METHODS: We identified 5418 ultrasound-guided thyroid nodule FNAs performed in our radiology department from 2004 through 2012. For each FNA, we recorded if the FNA was performed by a resident and if so the name of the resident and supervising attending radiologist. For each resident, we determined the level of training based on their graduation year from our residency program and the date of the FNA as well as prior surgical training and if they completed subsequent interventional radiology fellowship. Pathology reports were reviewed, and FNAs were classified as diagnostic or nondiagnostic (ND). Generalized mixed models were used to assess ND rate with postgraduate years, including residents with and without prior surgical training or if they subsequently completed an interventional radiology fellowship. RESULTS: Of the 5418 thyroid FNAs, 3164 (58.4%) were performed by a radiology resident under the direct supervision of an attending physician. There was a significant decrease in ND rate as postgraduate years increased (P < .05). A significant decrease in ND rate was found as postgraduate years increased for residents without prior surgical training (P = .0007) or subsequent training in interventional radiology (P = .0014); however, no significant decrease was found for residents with surgical training (P = .37) or completing an interventional radiology fellowship (P = .08). In addition, no significant difference was found for ND rate between postgraduate year 4 (PGY4) and PGY5 (P > .05). CONCLUSIONS: ND thyroid FNA rates progressively decrease with training level, suggesting that early and continued participation in procedures throughout residency improves outcomes. This is particularly true for residents without prior surgical training or subsequent interventional radiology fellowship.

Management of nodules with initially nondiagnostic results of thyroid fine-needle aspiration: can we avoid repeat biopsy?

PURPOSE: To identify demographic and ultrasonographic (US) features associated with malignancy after initially nondiagnostic results of fine-needle aspiration (FNA) to help clarify the role of repeat FNA, surgical excision, or serial US in these nodules. MATERIALS AND METHODS: This study was HIPAA compliant and institutional review board approved; informed consent was waived. Thyroid nodules (n = 5349) that underwent US-guided FNA in 2004-2012 were identified; 393 were single nodules with nondiagnostic FNA results but adequate cytologic, surgical, or US follow-up. Demographic information and diameters and volume at US at first biopsy were modeled with malignancy as outcome through medical record review. Exact logistic regression was used to model malignancy outcomes, demographic comparisons with age were made (Student t test, Satterthwaite test), and proportion confidence intervals (CIs) were estimated (Clopper-Pearson method). RESULTS: Of 393 nodules with initially nondiagnostic results, nine malignancies (2.3%) were subsequently diagnosed with repeat FNA (n = 2, 0.5%) or surgical pathologic examination (n = 7, 1.8%), 330 (84.0%) were benign, and 54 (13.7%) were stable or decreased in size at serial US (mean follow-up, 3.0 years; median, 2.5 years; range, 1.0-7.8 years). Patients with malignancies were significantly older (mean age, 62.7 years; median, 64 years; range, 47-77 years) than those without (mean age, 55.4 years; median, 57 years; range, 12-94 years; P = .0392). Odds of malignancy were 4.2 times higher for men versus women (P = .045) and increased significantly for each 1-cm increase in anteroposterior, minimum, and mean nodule diameter (1.78, 2.10, and 1.96, respectively). In 393 nodules, no malignancies were detected in cystic or spongiform nodules (both, n = 11, 2.8%; 95% CI: 1.4%, 5.0%), nodules with eggshell calcifications (n = 9, 2.3%; 95% CI: 1.1%, 4.3%), or indeterminate echogenic foci (n = 39, 9.9%; 95% CI: 7.2%, 13.3%). CONCLUSION: Very few malignancies were diagnosed with repeat FNA following nondiagnostic FNA results (two of 336, 0.6%); therefore, clinical and US follow-up may be more appropriate than repeat FNA following nondiagnostic biopsy results.

The chaperone protein clusterin may serve as a cerebrospinal fluid biomarker for chronic spinal cord disorders in the dog.

Chronic spinal cord dysfunction occurs in dogs as a consequence of diverse aetiologies, including long-standing spinal cord compression and insidious neurodegenerative conditions. One such neurodegenerative condition is canine degenerative myelopathy (DM), which clinically is a challenge to differentiate from other chronic spinal cord conditions. Although the clinical diagnosis of DM can be strengthened by the identification of the Sod1 mutations that are observed in affected dogs, genetic analysis alone is insufficient to provide a definitive diagnosis. There is a requirement to identify biomarkers that can differentiate conditions with a similar clinical presentation, thus facilitating patient diagnostic and management strategies. A comparison of the cerebrospinal fluid (CSF) protein gel electrophoresis profile between idiopathic epilepsy (IE) and DM identified a protein band that was more prominent in DM. This band was subsequently found to contain a multifunctional protein clusterin (apolipoprotein J) that is protective against endoplasmic reticulum (ER) stress-mediated apoptosis, oxidative stress, and also serves as an extracellular chaperone influencing protein aggregation. Western blot analysis of CSF clusterin confirmed elevated levels in DM compared to IE (p < 0.05). Analysis of spinal cord tissue from DM and control material found that clusterin expression was evident in neurons and that the clusterin mRNA levels from tissue extracts were elevated in DM compared to the control. The plasma clusterin levels was comparable between these groups. However, a comparison of clusterin CSF levels in a number of neurological conditions found that clusterin was elevated in both DM and chronic intervertebral disc disease (cIVDD) but not in meningoencephalitis and IE. These findings indicate that clusterin may potentially serve as a marker for chronic spinal cord disease in the dog; however, additional markers are required to differentiate DM from a concurrent condition such as cIVDD.

A protocol for the management of canine cerebrospinal fluid for the proteomic assessment of putative biomarkers.

Cerebrospinal fluid (CSF) is a potential source for disease-specific biomarkers that may assist in the staging and determining the prognosis of neurodegenerative conditions in animals. However, the validity of such putative biomarkers may be influenced by pre-analytical variables, including the procedures adopted to collect and store the CSF. This study assessed the effect of three handling practices on the stability of a panel of CSF proteins: clusterin (also known as apolipoprotein J), haptoglobin, cystatin C, and transthyretin (TTR). The three handling procedures for canine CSF were mimicked in the laboratory as follows: (1) storage in a refrigerator overnight (4 °C for 18 h); (2) carrying a sample in the pocket of a clinician (37 °C for 4h); and (3) mailing a sample to a remote laboratory for analysis (room temp for 48 h). The impact of these three scenarios on the concentrations of the selected proteins was assessed using Western blotting and compared to an aliquot of CSF that had been kept frozen. The level of clusterin was significantly reduced following 48 h at room temperature (P<0.05), while the concentration of the dimeric form of TTR increased following this handling procedure and also when held at 37 °C for 4h. A reducing agent prevented this increase at 37 °C. In conclusion, exposing CSF samples to various environmental conditions can significantly alter their protein content, a factor that must be considered in studies assessing potential biomarkers in canine CSF.

Aortic aneurysm generation in mice with targeted deletion of integrin-linked kinase in vascular smooth muscle cells.

RATIONALE: Integrin-linked kinase (ILK) is located at focal adhesions and links the extracellular matrix (ECM) to the actin cytoskeleton via ß1- and ß3-integrins. ILK plays a role in the activation of kinases including protein kinase B/Akt and glycogen synthase kinase 3ß and regulates cell proliferation, motility, and survival. OBJECTIVE: To determine the function of ILK in vascular smooth muscle cells (SMCs) in vivo. METHODS AND RESULTS: SM22Cre(+)Ilk(Fl/Fl) conditional mutant mice were generated in which the Ilk gene was selectively ablated in SMCs. SM22Cre(+)Ilk(Fl/Fl) conditional mutant mice survive to birth but die in the perinatal period exhibiting multiple vascular pathologies including aneurysmal dilatation of the aorta and patent ductus arteriosus (PDA). Defects in morphogenetic development of the aorta were observed as early as E12.5 in SM22Cre(+)Ilk(Fl/Fl) mutant embryos. By late gestation (E16.5 to 18.5), striking expansion of the thoracic aorta was observed in ILK mutant embryos. Histological analyses revealed that the structural organization of the arterial tunica media is severely disrupted with profound derangements in SMC morphology, cell-cell, and cell-matrix relationships, including disruption of the elastic lamellae. ILK deletion in primary aortic SMCs results in alterations of RhoA/cytoskeletal signaling transduced through aberrant localization of myocardin-related transcription factor (MRTF)-A repressing the transcription and expression of SMC genes, which are required for the maintenance of the contractile SMC phenotype. CONCLUSIONS: These data identify a molecular pathway linking ILK signaling to the contractile SMC gene program. Activation of this pathway is required for morphogenetic development of the aorta and ductus arteriosus during embryonic and postnatal survival.

High-resolution diffusion tensor imaging of fixed brain in a mouse model of Pelizaeus-Merzbacher disease: comparison with quantitative measures of white matter pathology.

Diffusion tensor imaging (DTI) is a powerful technique for the noninvasive assessment of the central nervous system. To facilitate the application of this technique to in vivo studies, we characterised a mouse model of the leukodystrophy, Pelizaeus-Merzbacher disease (PMD), comparing high-resolution ex vivo DTI findings with quantitative histological analysis of selected areas of the brain. The mice used in this study (Plp1-transgenic) carry transgenic copies of the Plp1 gene and are models for PMD as a result of gene duplication. Plp1 transgenic mice display a mild ataxia and experience frequent seizures around the time at which they were imaged. Axial (λ(1) ) and radial (RD) diffusivities and fractional anisotropy (FA) data were analysed using an exploratory whole-brain voxel-based method, a voxel-based approach using tract-based spatial statistics (TBSS), and by application of conventional region of interest (ROI) analyses to selected white matter tracts. Raw t value maps and TBSS analyses indicated widespread changes throughout the brain of Plp1-transgenic mice compared with the wild-type. ROI analyses of the corpus callosum, anterior commissure and hippocampal fimbria showed that FA was reduced significantly, whereas λ(1) and RD were increased significantly, in Plp1-transgenic mice compared with the wild-type. The DTI data derived from ROI analyses were subsequently compared with histological measures taken in the same regions. These revealed an almost complete absence of myelin, preservation of axons, marked astrocytosis and increased or unchanged cell densities. These data contribute to our growing understanding of the basis of anisotropic water diffusion in the normal and diseased nervous system.

PLP/DM20 expression and turnover in a transgenic mouse model of Pelizaeus-Merzbacher disease.

The most common cause of Pelizaeus-Merzbacher (PMD) is due to duplication of the PLP1 gene but it is unclear how increased gene dosage affects PLP turnover and causes dysmyelination. We have studied the dynamics of PLP/DM20 in a transgenic mouse model of PMD with increased gene dosage of the proteolipid protein gene (Plp1). The turnover of PLP/DM20 were investigated using an ex-vivo brain slice system and cultured oligodendrocytes. Homozygous mice have reduced PLP translation, markedly enhanced PLP degradation, and markedly reduced incorporation of PLP into myelin. Proteasome inhibition (MG132) prevented the enhanced degradation. Numerous autophagic vesicles are present in homozygous transgenic mice that may influence protein dynamics. Surprisingly, promoting autophagy with rapamycin decreases the degradation of nascent PLP suggesting autophagic vacuoles serve as a cellular storage compartment. We suggest that there are multiple subcellular fates of PLP/DM20 when overexpressed: the vast majority being degraded by the proteasome, a proportion sequestered into autophagic vacuoles, probably fused with endolysosomes, and only a small proportion entering the myelin sheath, where its association with lipid rafts is perturbed. Transgenic oligodendrocytes have fewer membrane sheets and this phenotype is improved with siRNA-mediated knockdown of PLP expression that promotes the formation of MBP+ myelin-like sheets. This finding suggests that RNAi technology is in principle applicable to improve CNS myelination when compromised by PLP/DM20 overexpression.

Development of occupational exposure limits for the Hanford tank farms.

Production of plutonium for the United States' nuclear weapons program from the 1940s to the 1980s generated 53 million gallons of radioactive chemical waste, which is stored in 177 underground tanks at the Hanford site in southeastern Washington State. Recent attempts to begin the retrieval and treatment of these wastes require moving the waste to more modern tanks and result in potential exposure of the workers to unfamiliar odors emanating from headspace in the tanks. Given the unknown risks involved, workers were placed on supplied air respiratory protection. CH2MHILL, the managers of the Hanford site tank farms, asked an Independent Toxicology Panel (ITP) to assist them in issues relating to an industrial hygiene and risk assessment problem. The ITP was called upon to help determine the risk of exposure to vapors from the tanks, and in general develop a strategy for solution of the problem. This paper presents the methods used to determine the chemicals of potential concern (COPCs) and the resultant development of screening values and Acceptable Occupational Exposure Limits (AOELs) for these COPCs. A total of 1826 chemicals were inventoried and evaluated. Over 1500 chemicals were identified in the waste tanks headspaces and more than 600 of these were assigned screening values; 72 of these compounds were recommended for AOEL development. Included in this list of 72 were 57 COPCs identified by the ITP and of these 47 were subsequently assigned AOELs. An exhaustive exposure assessment strategy was developed by the CH2MHILL industrial hygiene department to evaluate these COPCs.

Modulation of rumpshaker phenotype with wild-type PLP/DM20 suggests several pathogenic mechanisms.

The rumpshaker mutation of the murine myelin proteolipid protein 1 (Plp1) gene generates misfolded PLP/DM20 protein, resulting in dysmyelination, increased oligodendrocyte apoptosis, and death prior to P40 when expressed on the C57 BL/6 background. In this study, we used transgenic complementation to normalize the levels of PLP/DM20 in myelin with wild-type protein to determine whether loss of normal PLP function or gain of toxic function is responsible for dysmyelination in the rumpshaker. Restoring myelin PLP/DM20 levels extended the survival time to at least P60, significantly reduced the density of apoptotic cells, increased myelin volume, and restored normal periodicity of myelin. Biochemical analysis found that several myelin proteins that are reduced in rumpshaker, including MAG, CNP, and SirT2, are markedly elevated at peak myelination (P20) in the rumpshaker transgenic mouse. Myelin basic protein, however, remained low at peak myelination but was restored at P60 when myelin had matured and entered into a maintenance phase. Markers of the unfolded protein response (UPR), BiP and XBP1, remained activated with the introduction of wild-type PLP. These data demonstrate that restoring wild-type PLP/DM20 levels in rumpshaker improves the phenotype and the integrity of myelin, but hypomyelination persists and stress pathways remain activated. This suggests that both gain- and loss-of-function mechanisms are involved in the pathogenesis of the rumpshaker.

Pharmacological inhibition of BLT1 diminishes early abdominal aneurysm formation.

Leukotriene B(4) (LTB(4)) is a pro-inflammatory lipid mediator generated by the enzymes 5-lipoxygenase (5-LO) and LTA(4)-hydrolase. LTB(4) signals primarily through its G protein-coupled receptor BLT1, which is highly expressed on specific leukocyte subsets. Recent genetic studies in humans as well as knockout studies in mice have implicated the leukotriene synthesis pathway in several vascular pathologies. Here we tested the hypothesis that pharmacological inhibition of BLT1 diminishes abdominal aortic aneurysm (AAA) formation, a major complication associated with atherosclerotic vascular disease. Chow-fed Apoe(-/-) mice were treated with a 4-week infusion of Angiotensin II (AngII, 1000 ng/(kg min)) beginning at 10 weeks of age, in a well-established murine AAA model. Administration of the selective BLT1 antagonist CP-105,696 beginning simultaneously with AngII infusion reduced the incidence of AAA formation from 82% to 40% (p<0.05). There was a concordant reduction in maximal aortic diameter from 2.35 mm to 1.56 mm (p<0.05). While administration of the antagonist on day 14 after the onset of AngII infusion diminished lesional macrophage accumulation, it did not significantly alter the size of AAA by day 42. Thus, pharmacological inhibition of BLT1 may ultimately hold clinical promise, but early intervention may be critical.

Kynurenic acid triggers firm arrest of leukocytes to vascular endothelium under flow conditions.

Recent studies have demonstrated that kynurenic acid (KYNA), a compound produced endogenously by the interferon-gamma-induced degradation of tryptophan by indoleamine 2,3-dioxygenase, activates the previously orphaned G protein-coupled receptor, GPR35. This receptor is expressed in immune tissues, although its potential function in immunomodulation remains to be explored. We determined that GPR35 was most highly expressed on human peripheral monocytes. In an in vitro vascular flow model, KYNA triggered the firm arrest of monocytes to both fibronectin and ICAM-1, via beta(1) integrin- and beta(2) integrin-mediated mechanisms, respectively. Incubation of monocytes with pertussis toxin prior to use in flow experiments significantly reduced the KYNA-induced monocyte adhesion, suggesting that adhesion is triggered by a G(i)-mediated process. Furthermore, KYNA-triggered adhesion of monocytic cells was reduced by short hairpin RNA-mediated silencing of GPR35. Although GPR35 is expressed at slightly lower levels on neutrophils, KYNA induced firm adhesion of these cells to an ICAM-1-expressing monolayer as well. KYNA also elicited neutrophil shedding of surface L-selectin, another indicator of leukocyte activation. Taken together, these data suggest that KYNA could be an important early mediator of leukocyte recruitment.

Isolation of rare cells from cell mixtures by dielectrophoresis.

The application of dielectrophoretic field-flow fractionation (depFFF) to the isolation of circulating tumor cells (CTCs) from clinical blood specimens was studied using simulated cell mixtures of three different cultured tumor cell types with peripheral blood. The depFFF method can not only exploit intrinsic tumor cell properties so that labeling is unnecessary but can also deliver unmodified, viable tumor cells for culture and/or all types of molecular analysis. We investigated tumor cell recovery efficiency as a function of cell loading for a 25 mm wide x 300 mm long depFFF chamber. More than 90% of tumor cells were recovered for small samples but a larger chamber will be required if similarly high recovery efficiencies are to be realized for 10 mL blood specimens used CTC analysis in clinics. We show that the factor limiting isolation efficiency is cell-cell dielectric interactions and that isolation protocols should be completed within approximately 15 min in order to avoid changes in cell dielectric properties associated with ion leakage.

Gene expression profiles differentiating between breast cancers clinically responsive or resistant to letrozole.

PURPOSE: Endocrine agents, such as letrozole, are established in the treatment of hormone-dependent breast cancer. However, response rates are only 50% to 70% in the neoadjuvant setting and lower in advanced disease. Thus there is a need to identify novel markers predicting for response and to understand molecular mechanisms of resistance. PATIENTS AND METHODS: Sequential tumor biopsies were taken before and after 10 to 14 days of neoadjuvant treatment with letrozole in patients with estrogen receptor-rich breast cancer. Expression profiles on high-density microarray chips were then related to clinical responses as assessed from tumor volume measurements after 3 months of treatment. RESULTS: Of 52 patients, 37 (71%) were classified as having a clinical response to letrozole and 15 being clinically resistant. Bioinformatic analysis identified 205 covariables (69 baseline expression, 45 day 14 expression, and 91 change in expression with treatment) which differentiated between clinical responders and nonresponders. Hierarchical clustering using the variables separated responders and nonresponders into two distinct groups. Ontological assessment indicated that discriminating genes were enriched toward cellular biosynthetic processes. In particular, functional gene assessment showed ribosomal protein probes to have higher baseline expression in tumors responsive to letrozole and increased expression with treatment in nonresponding cases. CONCLUSION: To our knowledge, this is the first study to describe genetic covariables and molecular processes discriminating between tumors clinically responsive and resistant to an aromatase inhibitor. The understanding of such molecular phenotypes will be important in optimizing the clinical use of aromatase inhibitors, both in terms of identifying responsive breast cancers and developing new agents to target resistance pathways.

Interferon-gamma and the interferon-inducible chemokine CXCL10 protect against aneurysm formation and rupture.

BACKGROUND: Vascular disease can manifest as stenotic plaques or ectatic aneurysms, although the mechanisms culminating in these divergent disease manifestations remain poorly understood. T-helper type 1 cytokines, including interferon-gamma and CXCL10, have been strongly implicated in atherosclerotic plaque development. METHODS AND RESULTS: Here, we specifically examined their role in the formation of abdominal aortic aneurysms in the angiotensin II-induced murine model. Unexpectedly, we found increased suprarenal aortic diameters, abdominal aortic aneurysm incidence, and aneurysmal death in apolipoprotein E- and interferon-gamma-deficient (Apoe(-/-)/Ifng(-/-)) mice compared with Apoe(-/-) controls, although atherosclerotic luminal plaque formation was attenuated. The interferon-gamma-inducible T-cell chemoattractant CXCL10 was highly induced by angiotensin II infusion in Apoe(-/-) mice, but this induction was markedly attenuated in Apoe(-/-)/Ifng(-/-) mice. Apoe(-/-)/Cxcl10(-/-) mice had decreased luminal plaque but also increased aortic size, worse morphological grades of aneurysms, and a higher incidence of death due to aortic rupture than Apoe(-/-) controls. Furthermore, abdominal aortic aneurysms in Apoe(-/-)/Cxcl10(-/-) mice were enriched for non-T-helper type 1-related signals, including transforming growth factor-beta1. Treatment of Apoe(-/-)/Cxcl10(-/-) mice with anti-transforming growth factor-beta neutralizing antibody diminished angiotensin II-induced aortic dilation. CONCLUSIONS: The present study defines a novel pathway in which interferon-gamma and its effector, CXCL10, contribute to divergent pathways in abdominal aortic aneurysm versus plaque formation, inhibiting the former pathology but promoting the latter. Thus, efforts to develop antiinflammatory strategies for atherosclerosis must carefully consider potential effects on all manifestations of vascular disease.